Methods and compositions for stimulating collagen synthesis in skin cells

ABSTRACT

A composition comprising a first peptide selected from tripeptide, tetrapeptide and mixtures thereof; a second peptide selected from pentapeptide, hexapeptide, and mixtures thereof; an extract from the Laminaria genus, and whey protein, and a method for stimulating collagen synthesis in skin cells.

TECHNICAL FIELD

The invention is in the field of compositions for treating aging skin,and with particular efficacy in stimulating synthesis of collagen inskin cells.

BACKGROUND OF THE INVENTION

Collagen is one of the main structural proteins in skin. It can be foundin the fibrillar or non-fibrillar form. The fibrillar form is mostcommon and includes collagen subtypes I, II, III, V, and XI. Types I,IV, and V are most often associated with skin and dermal tissue.Collagen found in the skin typically diminishes with age and causeslaxity, lines, and wrinkles on skin. Any active ingredient that inducesskin cells to increase collagen synthesis is desirable because itameliorates the adverse effects of collagen deficiency in skin cellswhich causes lines, wrinkles, and skin laxity.

It has been discovered that certain tripeptides or tetrapeptides have asynergistic effect in stimulating collagen synthesis in skin cells whencombined with a combination of at least one pentapeptide or hexapeptide,at least one extract from Laminaria genus, and whey protein.

SUMMARY OF THE INVENTION

The invention is directed to a topical composition comprising a firstpeptide selected from a tripeptide, tetrapeptide or mixtures thereof; asecond peptide selected from the group consisting of a pentapeptide,hexapeptide, or mixtures thereof, at least one extract from theLaminaria genus, and whey protein.

The invention is also directed to a method for stimulating collagensynthesis in skin cells by topically applying a composition comprisingat least one first peptide selected from a tripeptide, tetrapeptide, ormixtures thereof; at least one second peptide selected from apentapeptide, hexapeptide, or mixtures thereof, at least one extractfrom Laminaria genus, and whey protein.

DETAILED DESCRIPTION

The composition of the invention may be in the liquid, semi-solid, orsolid form, and may be in the emulsion, solution, suspension, oranhydrous form. If in the solution or suspension form, the compositionmay contain from about 50 to 99.9% water. If in the emulsion form, thecomposition may contain from about 5-95% water and from about 5-95% oil.If in the anhydrous form, the composition may comprise from about 10-99%oil and 10-99% solidifying agents.

I. The Tripeptide or Tetrapeptide

The composition of the invention contains at least one tripeptide ortetrapeptide, or mixtures thereof. Suggested ranges are from about0.000001 to about 10%, preferably from about 0.000005 to 5%, morepreferably from about 0.00001 to 2.5%.

A particularly preferred tripeptide has the INCI name Tripeptide-32 andhas the following amino acid sequence:

S-T-P-NH₂ Ser-Thr-Pro-NH₂

A particularly preferred tetrapeptide has the INCI name Tetrapeptide-26which has the amino acid sequence:

(SEQ ID No. 1) S-P-L-Q-NH₂ Ser-Pro-Leu-Gln-NH₂

These peptides are manufactured by ISP-Vinscience under the trademarkChronolux® having the INCI name Tripeptide-32 or Chronogen® having theINCI name Tetrapeptide-26.

The Pentapeptide or Hexapeptide

The composition contains at least one pentapeptide or hexapeptide ormixtures thereof in an amount ranging from about 0.000001 to 5%,preferably from about 0.00001 to 2%, more preferably from about 0.0005to 1% by weight of the total composition.

The pentapeptide or hexapeptide may be substituted with acyl groups suchas acetyl, palmitoyl, myristoyl and the like. Further specific examplesinclude hexapeptides 1-60, said range including each whole integerbetween 1 and 60, hexapeptides that are acetylated, palmitoylated ormyristoylated such as acetyl hexapeptides 1, 7, 8, 19, 20, 22, 24, 30,31, 37, 38, 39, or 40. Particularly preferred is Acetyl Hexapeptide-8which is obtained by the acetylation of Hexapeptide-8, a syntheticpeptide containing arginine, glutamic acid, glutamine, and methionine.Acetyl Hexpeptide-8, represented by sequence listing

(SEQ ID No. 2) Ac-E-E-M-Q-R-R-NH₂. Ac-Glu-Glu-Met-Gln-Arg-Arg-NH₂can be purchased from Lipotec S.A. under the trade name Argireline®,which is a solution of about 0.05 parts Acetyl Hexpeptide-8, 99.35 partswater, with the remainder preservatives.

Also suitable are pentapeptides which may be acetylated, palmitoylated,or myristoylated. Examples of such pentapeptides include Pentapepides1-50 which includes each integer inbetween. Particularly preferred isPalmitoyl pentapeptide-5.

Particularly preferred are oligopeptides having the INCI namesAcetyl-hexapeptide-8, Palmitoyl hexapeptide-12, Pentapeptide-3,Palmitoyl pentapeptide-5 or combinations thereof.

These peptides are defined by the International Nomenclature forCosmetic Ingredients (INCI) and are terms known in the art.

The Extract from Laminaria Genus

The composition contains at least one extract from the Laminaria genus.Laminaria is a genus that contains 30+ species of the brown algaePhaeophyceae, often referred to as kelp. Such extracts from theLaminaria genus include those of species abyssalis, agardhii,appressirhiza, brasiliensis, brongardiana, bulbosa, bullata, complanata,digitata, ephemera, farlowii, groenlandica, hyperborea, inclinitorhiza,multiplicata, nigripes, ochroleuca, pallida, platymeris, rodriguezi,ruprechtii, sachalinensis, setchellii, sinclairii, solidungula, oryezoensis. Preferred is where the extract from the Laminaria genus isalso an activator of Sirtuin 3. Preferred is where the extract is fromLaminaria digitata, and more specifically an extract having laminarincontent and/or a mannitol content ranging from 0.5 to 3% by weight, orfrom about 0.75 to 2.5%, by weight, or most preferably from about 1% byweight or greater, preferably around 2%. An example of a suitableextract of Laminaria digitata may be purchased from Barnet Productsunder the tradename Mitostime Di which is a mixture of 91 parts water, 8parts Laminaria digitata extract, and 1 part preservative. Preferablythe Laminaria digitata extract is obtained by aqueous extraction andleaching of lyophilized algae and sterilizing the microfiltration,followed by reverse osmosis to concentrate the active molecules.

In the preferred embodiment of the invention the Laminaria extract maybe present in the composition in amounts ranging from 0.0001 to 5%,preferably from about 0.001 to 2.5%, more preferably from about 0.01 to1%.

Whey Protein

The composition contains whey protein, in an amount ranging from 0.01 to5%, preferably from about 0.05 to 3%, more preferably from about 0.1 to2% by weight of the total composition.

Whey protein is the polypeptide obtained from the fluid part of milkafter separation from curds. The whey protein may be hydrolyzed. Mostpreferred is a whey protein sold by Glanbia Foods having the trade namewhey protein NXP.

In one embodiment, the pentapeptide or hexapeptide, Laminaria extractand whey protein may be supplied to the composition in the form of apre-blend that can then be formulated into the final product. In thiscase a ratio of from about 2-20 parts of pentapeptide or hexapeptide,1-10 parts Laminaria extract, and 0.1 to 5 parts whey protein isappropriate. Most preferred is a ratio of 10 parts Acetyl hexapeptide-8,5 parts Laminaria digitata extract, and 1 part whey protein.

The composition of the invention, which is a collagen synthesisstimulating composition, may consist of the first peptide which is thetri- or tetrapeptide or mixtures thereof, the second peptide which isthe penta- or hexapeptide or mixtures thereof, the extract from theLaminaria genus, and whey protein and no other ingredients.

The composition of the invention may also “consist essentially of” thefirst peptide which is the tri- or tetrapeptide; the second peptidewhich is the penta- or hexapeptide, the extract from the Laminariagenus, and whey protein, which means a composition that contains thefour ingredients and only additional ingredients that do not affect thatbasic and novel characteristics of the composition such as water,preservatives, antioxidants, pH adjusters, solvents, and inertingredients such a silicones that do not affect the collagen stimulatingactivity of the composition.

The composition of the invention may also “comprise” the fouringredients mentioned and include other ingredients including but notlimited to those set forth herein.

Other Ingredients

Oils

Suitable oils include silicones, esters, vegetable oils, synthetic oils,including but not limited to those set forth herein. The oils may bevolatile or nonvolatile, and are preferably in the form of a pourableliquid at room temperature. If present, the oils may range from about0.5 to 85%, preferably from about 1-75%, more preferably from about5-65% by weight of the total composition.

Cyclic and linear volatile silicones are available from variouscommercial sources including Dow Corning Corporation and GeneralElectric. The Dow Corning linear volatile silicones are sold under thetrade names Dow Corning 244, 245, 344, and 200 fluids. These fluidsinclude hexamethyldisiloxane (viscosity 0.65 centistokes (abbreviatedcst)), octamethyltrisiloxane (1.0 cst), decamethyltetrasiloxane (1.5cst), dodecamethylpentasiloxane (2 cst) and mixtures thereof, with allviscosity measurements being at 25° C.

Suitable branched volatile silicones include alkyl trimethicones such asmethyl trimethicone, a branched volatile silicone having the generalformula:

Methyl trimethicone may be p

purchased from Shin-Etsu Silicones under the trade name TMF-1.5, havinga viscosity of 1.5 centistokes at 25° C.

Also suitable are various straight or branched chain paraffinichydrocarbons having 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18,19, or 20 carbon atoms, more preferably 8 to 16 carbon atoms. Suitablehydrocarbons include pentane, hexane, heptane, decane, dodecane,tetradecane, tridecane, and C₈₋₂₀ isoparaffins. Suitable C₁₂isoparaffins are manufactured by Permethyl Corporation under thetradename Permethyl 99A. Various C₁₆ isoparaffins commerciallyavailable, such as isohexadecane (having the tradename Permethyl R), arealso suitable.

Also suitable are esters formed by the reaction of a carboxylic acid andan alcohol. The alcohol and the carboxylic acids may both have fatty(C6-30) chains. Examples include hexyl laurate, butyl isostearate,hexadecyl isostearate, cetyl palmitate, isostearyl neopentanoate,stearyl heptanoate, isostearyl isononanoate, stearyl lactate, stearyloctanoate, stearyl stearate, isononyl isononanoate, and so on.

The ester may also be in the dimer or trimer form. Examples of suchesters include diisotearyl malate, neopentyl glycol dioctanoate, dibutylsebacate, dicetearyl dimer dilinoleate, dicetyl adipate, diisocetyladipate, diisononyl adipate, diisostearyl dimer dilinoleate,diisostearyl fumarate, diisostearyl malate, dioctyl malate, and so on.

Examples of other types of esters include those from arachidonic,citric, or behenic acids, such as triarachidin, tributyl citrate,triisostearyl citrate, tri C₁₂₋₁₃ alkyl citrate, tricaprylin,tricaprylyl citrate, tridecyl behenate, trioctyldodecyl citrate,tridecyl behenate; or tridecyl cocoate, tridecyl isononanoate, and soon.

Synthetic or naturally occurring glyceryl esters of fatty acids, ortriglycerides, are also suitable for use in the compositions. Bothvegetable and animal sources may be used. Examples of such oils includecastor oil, lanolin oil, C₁₀₋₁₈ triglycerides,caprylic/capric/triglycerides, sweet almond oil, apricot kernel oil,sesame oil, camelina sativa oil, tamanu seed oil, coconut oil, corn oil,cottonseed oil, linseed oil, ink oil, olive oil, palm oil, illipebutter, rapeseed oil, soybean oil, grapeseed oil, sunflower seed oil,walnut oil, and the like.

Also suitable are synthetic or semi-synthetic glyceryl esters, such asfatty acid mono-, di-, and triglycerides which are natural fats or oilsthat have been modified, for example, mono-, di- or triesters of polyolssuch as glycerin. In an example, a fatty (C₁₂₋₂₂) carboxylic acid isreacted with one or more repeating glyceryl groups. glyceryl stearate,diglyceryl diiosostearate, polyglyceryl-3 isostearate, polyglyceryl-4isostearate, polyglyceryl-6 ricinoleate, glyceryl dioleate, glyceryldiisotearate, glyceryl tetraisostearate, glyceryl trioctanoate,diglyceryl distearate, glyceryl linoleate, glyceryl myristate, glycerylisostearate, PEG castor oils, PEG glyceryl oleates, PEG glycerylstearates, PEG glyceryl tallowates, and so on.

Nonvolatile silicone oils, both water soluble and water insoluble, arealso suitable for use in the composition. Such silicones preferably havea viscosity ranging from about greater than 5 to 800,000 cst, preferably20 to 200,000 cst at 25° C. Suitable water insoluble silicones includeamine functional silicones such as amodimethicone. Examples includedimethicone, phenyl dimethicone, diphenyl dimethicone, phenyltrimethicone, or trimethylsiloxyphenyl dimethicone. Other examplesinclude alkyl dimethicones such as cetyl dimethicone, stearyldimethcone, behenyl dimethicone, and the like.

Surfactants

The composition may contain one or more surfactants, especially if inthe emulsion form. However, such surfactants may be used if thecompositions are anhydrous also, and will assist in dispersingingredients that have polarity, for example pigments. Such surfactantsmay be silicone or organic based. The surfactants will aid in theformation of stable emulsions of either the water-in-oil or oil-in-waterform. If present, the surfactant may range from about 0.001 to 30%,preferably from about 0.005 to 25%, more preferably from about 0.1 to20% by weight of the total composition.

Silicone surfactants may be generically referred to as dimethiconecopolyol or alkyl dimethicone copolyol. In some cases the number ofrepeating ethylene oxide or propylene oxide units in the polymer arealso specified, such as a dimethicone copolyol that is also referred toas PEG-15/PPG-10 dimethicone, which refers to a dimethicone havingsubstituents containing 15 ethylene glycol units and 10 propylene glycolunits on the siloxane backbone. It is also possible for one or more ofthe methyl groups in the above general structure to be substituted witha longer chain alkyl (e.g. ethyl, propyl, butyl, etc.) or an ether suchas methyl ether, ethyl ether, propyl ether, butyl ether, and the like.

Examples of silicone surfactants are those sold by Dow Corning under thetradename Dow Corning 3225C Formulation Aid having the CTFA namecyclotetrasiloxane (and) cyclopentasiloxane (and) PEG/PPG-18dimethicone; or 5225C Formulation Aid, having the CTFA namecyclopentasiloxane (and) PEG/PPG-18/18 dimethicone; or Dow Coming 190Surfactant having the CTFA name PEG/PPG-18/18 dimethicone; or DowCorning 193 Fluid, Dow Corning 5200 having the CTFA name laurylPEG/PPG-18/18 methicone; or Abil EM 90 having the CTFA name cetylPEG/PPG-14/14 dimethicone sold by Goldschmidt; or Abil EM 97 having theCTFA name bis-cetyl PEG/PPG-14/14 dimethicone sold by Goldschmidt; orAbil WE 09 having the CTFA name cetyl PEG/PPG-10/1 dimethicone in amixture also containing polyglyceryl-4 isostearate and hexyl laurate; orKF-6011 sold by Shin-Etsu Silicones having the CTFA name PEG-11 methylether dimethicone; KF-6012 sold by Shin-Etsu Silicones having the CTFAname PEG/PPG-20/22 butyl ether dimethicone; or KF-6013 sold by Shin-EtsuSilicones having the CTFA name PEG-9 dimethicone; or KF-6015 sold byShin-Etsu Silicones having the CTFA name PEG-3 dimethicone; or KF-6016sold by Shin-Etsu Silicones having the CTFA name PEG-9 methyl etherdimethicone; or KF-6017 sold by Shin-Etsu Silicones having the CTFA namePEG-10 dimethicone; or KF-6038 sold by Shin-Etsu Silicones having theCTFA name lauryl PEG-9 polydimethylsiloxyethyl dimethicone.

Also suitable are various types of crosslinked silicone surfactants thatare often referred to as emulsifying elastomers that contain at leastone hydrophilic moiety such as polyoxyalkylenated groups.Polyoxyalkylenated silicone elastomers that may be used in at least oneembodiment of the invention include those sold by Shin-Etsu Siliconesunder the names KSG-21, KSG-20, KSG-30, KSG-31, KSG-32, KSG-33; KSG-210which is dimethicone/PEG-10/15 crosspolymer dispersed in dimethicone;KSG-310 which is PEG-15 lauryl dimethicone crosspolymer; KSG-320 whichis PEG-15 lauryl dimethicone crosspolymer dispersed in isododecane;KSG-330 (the former dispersed in triethylhexanoin), KSG-340 which is amixture of PEG-10 lauryl dimethicone crosspolymer and PEG-15 lauryldimethicone crosspolymer.

Also suitable are polyglycerolated silicone elastomers like thosedisclosed in PCT/WO 2004/024798, which is hereby incorporated byreference in its entirety. Such elastomers include Shin-Etsu's KSGseries, such as KSG-710 which is dimethicone/polyglycerin-3 crosspolymerdispersed in dimethicone; or lauryl dimethicone/polyglycerin-3crosspolymer dispersed in a variety of solvent such as isododecane,dimethicone, triethylhexanoin, sold under the Shin-Etsu tradenamesKSG-810, KSG-820, KSG-830, or KSG-840. Also suitable are silicones soldby Dow Corning under the tradenames 9010 and DC9011.

The composition may comprise one or more nonionic organic surfactants.Suitable nonionic surfactants include alkoxylated alcohols, or ethers,formed by the reaction of an alcohol with an alkylene oxide, usuallyethylene or propylene oxide. Preferably the alcohol is either a fattyalcohol having 6 to 30 carbon atoms. Examples of such ingredientsinclude Steareth 2-100, which is formed by the reaction of stearylalcohol and ethylene oxide and the number of ethylene oxide units rangesfrom 2 to 100; Beheneth 5-30 which is formed by the reaction of behenylalcohol and ethylene oxide where the number of repeating ethylene oxideunits is 5 to 30; Ceteareth 2-100, formed by the reaction of a mixtureof cetyl and stearyl alcohol with ethylene oxide, where the number ofrepeating ethylene oxide units in the molecule is 2 to 100; Ceteth 1-45which is formed by the reaction of cetyl alcohol and ethylene oxide, andthe number of repeating ethylene oxide units is 1 to 45, and so on. Allrecitations of units include all whole integers between the range.

Other alkoxylated alcohols are formed by the reaction of fatty acids andmono-, di- or polyhydric alcohols with an alkylene oxide. For example,the reaction products of C₆₋₃₀ fatty carboxylic acids and polyhydricalcohols which are monosaccharides such as glucose, galactose, methylglucose, and the like, with an alkoxylated alcohol. Examples includepolymeric alkylene glycols reacted with glyceryl fatty acid esters suchas PEG glyceryl oleates, PEG glyceryl stearate; or PEGpolyhydroxyalkanotes such as PEG dipolyhydroxystearate wherein thenumber of repeating ethylene glycol units ranges from 3 to 1000.

Other suitable nonionic surfactants include alkoxylated sorbitan andalkoxylated sorbitan derivatives. For example, alkoxylation, inparticular ethoxylation of sorbitan provides polyalkoxylated sorbitanderivatives. Esterification of polyalkoxylated sorbitan providessorbitan esters such as the polysorbates. For example, thepolyalkyoxylated sorbitan can be esterified with C6-30, preferablyC12-22 fatty acids. Examples of such ingredients include Polysorbates20-85, sorbitan oleate, sorbitan sesquioleate, sorbitan palmitate,sorbitan sesquiisostearate, sorbitan stearate, and so on.

Humectants

It may also be desirable to include one or more humectants in thecomposition. If present, such humectants may range from about 0.001 to25%, preferably from about 0.005 to 20%, more preferably from about 0.1to 15% by weight of the total composition. Examples of suitablehumectants include glycols, sugars, and the like. Suitable glycols arein monomeric or polymeric form and include polyethylene andpolypropylene glycols such as PEG 4-200, which are polyethylene glycolshaving from 4 to 200 repeating ethylene oxide units; as well as C₁₋₆alkylene glycols such as propylene glycol, butylene glycol, pentyleneglycol, and the like. Suitable sugars, some of which are also polyhydricalcohols, are also suitable humectants. Examples of such sugars includeglucose, fructose, honey, hydrogenated honey, inositol, maltose,mannitol, maltitol, sorbitol, sucrose, xylitol, xylose, and so on. Alsosuitable is urea. Preferably, the humectants used in the composition ofthe invention are C₁₋₆, preferably C₂₋₄ alkylene glycols, mostparticularly butylene glycol.

Botanical Extracts

It may be desirable to include one or more botanical extracts in thecompositions. If so, suggested ranges are from about 0.0001 to 10%,preferably about 0.0005 to 8%, more preferably about 0.001 to 5% byweight of the total composition. Suitable botanical extracts includeextracts from plants (herbs, roots, flowers, fruits, seeds) such asflowers, fruits, vegetables, and so on, including yeast ferment extract,Padina Pavonica extract, thermus thermophilis ferment extract, camelinasativa seed oil, boswellia serrata extract, olive extract, AribodopsisThaliana extract, Acacia Dealbata extract, Acer Saccharinum (sugarmaple), acidopholus, acorus, aesculus, agaricus, agave, agrimonia,algae, aloe, citrus, brassica, cinnamon, orange, apple, blueberry,cranberry, peach, pear, lemon, lime, pea, seaweed, caffeine, green tea,chamomile, willowbark, mulberry, poppy, and those set forth on pages1646 through 1660 of the CTFA Cosmetic Ingredient Handbook, EighthEdition, Volume 2. Further specific examples include, but are notlimited to, Glycyrrhiza Glabra, Salix Nigra, Macrocycstis Pyrifera,Pyrus Malus, Saxifraga Sarmentosa, Vitis Vinifera, Morus Nigra,Scutellaria Baicalensis, Anthemis Nobilis, Salvia Sclarea, RosmarinusOfficianalis, Citrus Medica Limonum, Panax Ginseng, SiegesbeckiaOrientalis, Fructus Mume, Ascophyllum Nodosum, Bifida Ferment lysate,Glycine Soja extract, Beta Vulgaris, Haberlea Rhodopensis, PolygonumCuspidatum, Citrus Aurantium Dulcis, Vitis Vinifera, SelaginellaTamariscina, Humulus Lupulus, Citrus Reticulata Peel, Punica Granatum,Asparagopsis, Curcuma Longa, Menyanthes Trifoliata, Helianthus Annuus,Hordeum Vulgare, Cucumis Sativus, Evernia Prunastri, Evernia Furfuracea,and mixtures thereof.

Particulate Materials

The compositions of the invention may contain particulate materials inthe form of pigments, inert particulates, or mixtures thereof. Ifpresent, suggested ranges are from about 0.01-75%, preferably about0.5-70%, more preferably about 0.1-65% by weight of the totalcomposition. In the case where the composition may comprise mixtures ofpigments and powders, suitable ranges include about 0.01-75% pigment and0.1-75% powder, such weights by weight of the total composition.

The particulate matter may be colored or non-colored powders. Suitablenon-pigmented powders include bismuth oxychloride, titanated mica, fumedsilica, spherical silica, polymethylmethacrylate, micronized teflon,boron nitride, acrylate copolymers, aluminum silicate, aluminum starchoctenylsuccinate, bentonite, calcium silicate, cellulose, chalk, cornstarch, diatomaceous earth, fuller's earth, glyceryl starch, hectorite,hydrated silica, kaolin, magnesium aluminum silicate, magnesiumtrisilicate, maltodextrin, montmorillonite, microcrystalline cellulose,rice starch, silica, talc, mica, titanium dioxide, zinc laurate, zincmyristate, zinc rosinate, alumina, attapulgite, calcium carbonate,calcium silicate, dextran, kaolin, nylon, silica silylate, silk powder,sericite, soy flour, tin oxide, titanium hydroxide, trimagnesiumphosphate, walnut shell powder, or mixtures thereof. The above mentionedpowders may be surface treated with lecithin, amino acids, mineral oil,silicone, or various other agents either alone or in combination, whichcoat the powder surface and render the particles more lipophilic innature.

Suitable pigments are organic or inorganic. Organic pigments aregenerally various aromatic types including azo, indigoid,triphenylmethane, anthroquinone, and xanthine dyes which are designatedas D&C and FD&C blues, browns, greens, oranges, reds, yellows, etc.Organic pigments generally consist of insoluble metallic salts ofcertified color additives, referred to as the Lakes. Inorganic pigmentsinclude iron oxides, ultramarines, chromium, chromium hydroxide colors,and mixtures thereof. Iron oxides of red, blue, yellow, brown, black,and mixtures thereof are suitable.

Vitamins and Antioxidants

The compositions of the invention may contain vitamins and/or coenzymes,as well as antioxidants. If so, 0.001-10%, preferably 0.01-8%, morepreferably 0.05-5% by weight of the total composition is suggested.Suitable vitamins include ascorbic acid and derivatives thereof such asascorbyl palmitate, tetrahexydecyl ascorbate, and so on; the B vitaminssuch as thiamine, riboflavin, pyridoxin, and so on, as well as coenzymessuch as thiamine pyrophoshate, flavin adenin dinucleotide, folic acid,pyridoxal phosphate, tetrahydrofolic acid, and so on. Also Vitamin A andderivatives thereof are suitable. Examples are retinyl palmitate,retinol. retinoic acid, as well as Vitamin A in the form of betacarotene. Also suitable is Vitamin E and derivatives thereof such asVitamin E acetate, nicotinate, or other esters thereof. In addition,Vitamins D and K are suitable.

The invention further comprises treating skin to stimulate collagensynthesis by topically applying a composition tri- or tetrapeptide, atleast one penta- or hexapeptide, at least one extract from the Laminariagenus, and whey protein. The compositions may be applied in the formsmentioned herein, as part of skin care regimens. For example, thecomposition may be applied to the skin as a night cream or cream appliedto skin prior to a period of bodily rest such as a nap or sleep. Thecomposition may be applied two times a day, in the morning and in theevening after cleansing the skin. The composition may be applied to theskin over skin care products, in the form of foundations or other colorcosmetics.

The invention will be further described in connection with the followingexamples which are set forth for the purposes of illustration only.

Example 1

Tripeptide-32 was tested at various concentrations for its ability tostimulate collagen production in normal human dermal fibroblasts(“NHDF”) from a 62 year old donor in a collagen induction assay.Tripeptide-32 was tested at 1× (0.2%) 0.5× (0.1%) and 0.2× (0.04%)concentration diluted in DMEM (“Dulbecco's Modified Eagle Medium”)supplemented with 10% Hyclone® bovine calf serum and 1% Cellgro®penicillin-streptomycin solution (“DMEM”). All percentages are by weightof the total composition.

Cell Growth and Maintenance.

Aged NHDF were obtained from ZenBio. Fibroblasts were cultured in DMEM1× (Life Tech) supplemented with 10% bovine calf serum (“BCS”) (Hyclone)and 1% PenStrep (“PS”) solution (Cellgro). Cells were regularlymaintained; subcultured as needed.

Plating Cells.

Normal human dermal fibroblasts (NHDF, aged) were plated on a 96-wellplate in supplemented DMEM (as indicated above). All rows, except forRow A, were seeded with cells (Row A was left blank to allow forbackground subtraction). Plate was allowed to incubate at standardconditions (37° C., 5% CO₂, 95% humidity) overnight.

Preparation & Treatment of Cells.

The following treatments were created in full media (1% P/S and 10% BCSsupplemented) DMEM: (1) Laminaria digitata extract (0.5%); Acetylhexapeptide-8 solution (a mixture of 0.05% Acetyl hexapeptide-8, 99.35%water and the remainder preservatives) (10%); and whey protein (0.1%,solid) with the remainder DMEM (the “Mixture”); (2)Mixture+Tripeptide-32: Laminaria digitata extract (0.5%), Acetylhexpeptide-8 solution (10%), whey protein (0.1%) and Tripeptide-32(0.2%) in DMEM at 1× (0.2%), 0.5× (0.1%), and 0.2× (0.04%)concentration; (3) Tripeptide-32 alone at 1× (0.2%), 0.5× (0.1%) and0.2× (0.04%) concentration. Each treatment (200 ul/well) was added tothe corresponding well of the 96-well plate. Treated plate was allowedto incubate at standard conditions (37° C., 5% CO₂, 95% humidity) for 5days.

Viability Assay.

Following 5 days of incubation, supernatants were harvested and storedprior to collagen analysis. A 10% alamar blue (Life Tech) solution wascreated in warmed (37° C.) full media and assay was completed exactly asper manufacturer's (Life Tech) protocol. Alamar Blue results weredetermined using a plate reader. Data was analyzed using the SoftMax Prosoftware and Excel.

Assessment of Collagen Production.

Collagen production was assessed using the Pro-collagen Type I collagenEIA Kit (Takara) as per manufacturer's protocol, exactly as described.Plate was read using the Gemini M2E plate reader and results werecompared.

The results are set forth below and show that Tripeptide-32 alone showsdecreased collagen synthesis at 0.2× (0.04%), and nearly insignificantincrease in collagen synthesis at 0.5× (0.1%) and 1× (0.2%)concentrations. The Mixture of Acetyl hexapeptide-8, Laminaria digitataextract, and whey protein demonstrates increased collagen synthesis.However, there is a synergistic increase in collagen synthesis whenTripeptide-32 is combined with the Mixture.

% increase in Test Material Concentration collagen induction DMEM Neat(Control) 0 Tripeptide-32 0.2X −15.39 Tripeptide-32 0.5X +1.17Tripeptide-32 1.0X +3.27 Mixture alone 0.2X +129.50 Mixture alone 0.5X+165.08 Mixture alone 1.0X +200 Mixture + Tripeptide-32 0.2X +148.13Mixture + Tripeptide-32 0.5X +214.57 Mixture + Tripeptide-32 1.0X+267.14

In general it is seen that when Tripeptide-32 is combined with themixture of Acetyl hexapeptide-8, Laminaria digitata extract, and wheyprotein there is a synergistic increase in collagen synthesis in cellswhich is unexpected given that Tripeptide-32 alone causes decreasedcollagen synthesis, or at best very marginal insignificant increases incollagen synthesis. Thus, this combination stimulates collagen synthesisin skin cells.

Example 2

The combination of Tripeptide-32 and a mixture of Acetyl hexapeptide-8,Laminaria digitata extract and whey protein were tested for collagenstimulation in NHDF from a 62 year old donor. Tripeptide-32 at aconcentration of 0.2% was combined with a mixture of Acetylhexapeptide-8 solution (0.05% Acetyl hexapeptide-8, 99.35% water,remainder preservatives) (10%); 0.5% Laminaria digitata extractsolution, and 0.1% whey protein in Supplemented DMEM.

The tests were performed according to the method in Example 1. Theresults are set forth below:

Percent Increase in Collagen Test Material: Synthesis over Mixture Alone(%) 0.2X Mixture + 0.2X Tripeptide-32 +8.44 0.5X Mixture + 0.5XTripeptide-32 +18.67 1.0X Mixture + 1.0X Tripeptide-32 +22.38

The above results show that addition of increasing concentrations ofTripeptide-32 extract to increasing concentrations of the Mixtureprovided a dose response increase in collagen synthesis in fibroblasts.As noted in Example 1, Tripeptide-32 itself largely causes decreasedcollagen synthesis in fibroblasts, and exhibits no dose responserelationship to increasing concentrations.

Example 3

A skin care composition according to the invention was made as follows:

Ingredient % by weight Water QS100 Isononyl isononanoate 6.0 C12-20 acidPEG-8 ester 3.0 Glycerin 2.6 Dimethicone 1.5 Shea butter 1.5 Cetylalcohol 1.4 Butylene glycol 1.2 PEG-100 stearate 0.75 Acetyl glucosamine0.50 Sucrose 0.50 Preservatives 1.0 Ammonium acryloyldimethyl taurate0.35 Sorbitol 0.35 Pentylene glycol 0.25 Algae extract 0.25 Caffeine0.20 Carbomer 0.20 Potassium cetyl phosphate 0.20 Tocopheryl acetate0.20 Ethylhexylglycerin 0.15 Aquacell* 1.00 Acrylates/C10-30 alkylacrylates crosspolymer 0.11 Phytofix** 0.20 Whey protein 0.10 Glucose0.10 Isoceteth-20 0.10 Laminaria digitata extract 0.04 Acetylhexapeptide 8 0.01 Tripeptide-32 0.20 *Aquacell: a mixture of water,Citrullus lanatus (watermelon) fruit extract, Pyrus malus (apple) fruitextract, Lens esculenta (Lentil) fruit extract, sodium lactate, andsodium PCA. **Phytofix: a mixture of propylene glycol dicaprate,Helianthus annus (sunflower) seed cake, Hordeum vulgare (barley)extract, Cucumis sativus (cucumber) fruit extract.

While the invention has been described in connection with the preferredembodiment, it is not intended to limit the scope of the invention tothe particular form set forth but, on the contrary, it is intended tocover such alternatives, modifications, and equivalents as may beincluded within the spirit and scope of the invention as defined by theappended claims.

The invention claimed is:
 1. A composition comprising a first peptideselected from the group consisting of Tripeptide-32, Tetrapeptide-26,and mixtures thereof; a second peptide selected from the groupconsisting of a pentapeptide selected from the group consisting ofPentapeptide-3, Palmitoyl pentapeptide-5 and mixtures thereof, ahexapeptide selected from the group consisting of Palmitoylhexapeptide-12, Acetyl hexapeptide-8, and mixtures thereof; at least oneextract from the Laminaria genus; and whey protein.
 2. The compositionof claim 1 wherein the first peptide is Tetrapeptide-26.
 3. Thecomposition of claim 1 wherein the first peptide is Tripeptide-32 and ispresent at 0.000001 to about 10%.
 4. The composition of claim 1 whereinthe second peptide is a hexapeptide.
 5. The composition of claim 4wherein the hexapeptide is Palmitoyl hexapeptide-12.
 6. The compositionof claim 4 wherein the acetylated hexapeptide is Acetyl hexapeptide-8and is present at 0.000001 to 5%.
 7. The composition of claim 1 whereinthe first peptide is a tripeptide that is Tripeptide-32, the secondpeptide is a hexapeptide that is Acetyl hexpeptide-8, and the extractfrom the Laminaria genus is from Laminaria digitata.
 8. The compositionof claim 7 wherein the extract from Laminaria digitata has a laminarinor mannitol content of 0.5 to 3% by weight of the total extract.
 9. Thecomposition of claim 8 wherein the Laminaria digitata extract isobtained by aqueous extraction and leaching of lyophilized algae. 10.The composition of claim 8 wherein the Laminaria digitata extract ispresent from 0.0001 to 5%.
 11. A composition comprising, by weight ofthe total composition: 0.000001 to 10% Tripeptide-32, 0.000001 to 5%Acetyl hexapeptide-8, 0.0001 to 5% Laminaria digitata extract, 0.05 to3% whey protein.
 12. The composition of claim 11 wherein the Laminariadigitata extract has a laminarin or mannitol content ranging from 0.5 to3% by weight of the total extract.
 13. The composition of claim 12wherein the Laminaria digitata extract is obtained by aqueous extractionand leaching of lyophilized algae.
 14. The composition of claim 11 inthe form of a skin cream, lotion, foundation makeup, or gel.
 15. Amethod for stimulating collagen synthesis in skin cells by topicallyapplying the composition of claim
 1. 16. The method of claim 15 whereinthe composition is in the form of a skin cream or lotion.
 17. The methodof claim 16 wherein the composition is applied once or twice per day.18. The method of claim 16 wherein the first peptide is a tripeptidethat is Tripeptide-32, the second peptide is a peptide that isTetrapeptide-26, and the extract from the Laminaria genus is fromLaminaria digitata.
 19. The method of claim 18 wherein the Tripeptide-32is present at 0.000001 to 10%, the Acetyl hexapeptide-8 is present at0.000001 to 5%, the Laminaria digitata extract is present at 0.0001 to5%, and the whey protein is present at 0.05 to 3%.
 20. The method ofclaim 19 wherein the composition is applied at night prior to retiring.21. A collagen stimulating composition comprising Tripeptide-32 incombination with a mixture of Laminaria digitata extract, whey protein,and Acetyl hexapeptide-8.
 22. The collagen stimulating composition ofclaim 21 showing a greater than additive increase in collagen synthesiswhen compared to the collagen stimulating activity of Tripeptide-32alone and the mixture of Laminaria digitata extract, whey protein, andAcetyl hexapeptide-8 alone.
 23. The collagen stimulating composition ofclaim 21 wherein the collagen stimulating activity is characterized byshowing a synergistic increase in collagen synthesis when tested onfibroblasts in vitro.